Study of the rate and spectrum of spontaneous mutations

Mutations are the initial force responsible for all aspects of genetic variation, and are a central part to evolution in all organisms. Yet despite its importance, the previously high cost that is associated with surveying mutations at a genome-wide scale has limited the understanding of the mutation process in eukaryotes. However, recent high-throughput sequencing technology has greatly reduced the cost of surveying mutations. By applying high-throughput sequencing to mutation accumulation experiments, we have begun to characterize the genome-wide mutation spectrum of eukaryotes. Across all eukaryotes, we observe a biased rate of G/C-\u3e A/T mutations that exceeds the number of A/T-\u3eG/C mutations. This finding is consistent with spontaneous deamination of cytosine or methylated cytosine. Alternate forces such as selection or G/C biased gene conversion must be driving eukaryotic genomes toward a higher G/C composition than expected from mutation bias. In Paramecium tetraurelia, we observe a nuclear mutation rate ∼75 fold lower than previously expected. When the base substitution rate per generation is extrapolated to the rate per expressed sexual cycle, it is equivalent to that observed in multicellular species with comparable genome sizes. This suggests that natural selection operates at germline expression, and favors a minimum rate that opposes random genetic drift. Using a natural population of Daphnia pulex we catalogue simple sequence repeats (SSR) and determine average heterozygosity of each SSR type. We find that SSR heterozygosity is motif specific, and positively correlated with repeat number as well as motif length. We identify a motif-dependent end-nucleotide polymorphism bias. Our observations also confirm the high frequency of multiple unit variation at large microsatellite loci. We observe that structural variants in C. elegans and S. cerevisiae, which are ∼1000 fold larger than base substitution rates on a per nucleotide basis, occur on the same order of magnitude as base substitutions. The rate and direction of structural gains and losses differ between yeast and C. elegans, and we hypothesize that the rate of structural variants corresponds with the coding portion of the genome. We also confirm a high rate of gene inversion and gene loss in the life history of C. elegans

Percutaneous Transhepatic Cholangioscopic Intervention in the Management of Complete Membranous Occlusion of Bilioenteric Anastomosis: Report of Two Cases

Postoperative biliary stricture is a relatively rare but serious complication of biliary surgery. Although Rouxen-Y hepaticojejunostomy or choledochojejunostomy are well-established and fundamental therapeutic approaches, their postoperative morbidity and mortality rates have been reported to be up to 33% and 13%, respectively. Recent studies suggest that percutaneous transhepatic intervention is an effective and less invasive therapeutic modality compared with traditional surgical treatment. Compared with fluoroscopic intervention, percutaneous with cholangioscopy may be more useful in biliary strictures, as it can provide visual information regarding the stricture site. We recently experienced two cases complete membranous occlusion of the bilioenteric anastomosis and successfully treated both patients using percutaneous transhepatic cholangioscopy

A genome-wide view of Caenorhabditis elegans base-substitution mutation processes

Knowledge of mutation processes is central to understanding virtually all evolutionary phenomena and the underlying nature of genetic disorders and cancers. However, the limitations of standard molecular mutation detection methods have historically precluded a genome-wide understanding of mutation rates and spectra in the nuclear genomes of multicellular organisms. We applied two high-throughput DNA sequencing technologies to identify and characterize hundreds of spontaneously arising base-substitution mutations in 10 Caenorhabditis elegans mutation-accumulation (MA)-line nuclear genomes. C. elegans mutation rate estimates were similar to previous calculations based on smaller numbers of mutations. Mutations were distributed uniformly within and among chromosomes and were not associated with recombination rate variation in the MA lines, suggesting that intragenomic variation in genetic hitchhiking and/or background selection are primarily responsible for the chromosomal distribution patterns of polymorphic nucleotides in C. elegans natural populations. A strong mutational bias from G/C to A/T nucleotides was detected in the MA lines, implicating oxidative DNA damage as a major endogenous mutagenic force in C. elegans. The observed mutational bias also suggests that the C. elegans nuclear genome cannot be at equilibrium because of mutation alone. Transversions dominate the spectrum of spontaneous mutations observed here, whereas transitions dominate patterns of allegedly neutral polymorphism in natural populations of C. elegans and many other animal species; this observation challenges the assumption that natural patterns of molecular variation in noncoding regions of the nuclear genome accurately reflect underlying mutation processes

Association of Plasma Levels of Resistin with Subcutaneous Fat Mass and Markers of Inflammation but not with Metabolic Determinants or Insulin Resistance

The aim of the present study was to investigate the relationship of plasma resistin levels with determinants of the metabolic syndrome (MetS) and anthropometric parameters in healthy Korean subjects. Plasma resistin levels were determined in 276 subjects. In subjects with MetS, the plasma resistin levels were not significantly increased compared to those without MetS (8.3±4.3 ng/mL vs. 8.5±3.6 ng/mL, respectively, P=0.84). In addition, the plasma resistin levels were not correlated with the body mass index, the waist circumference, homeostasis model assessment-insulin resistance (HOMA-IR), fasting plasma glucose or insulin levels. However, the plasma resistin levels were positively correlated with the abdominal subcutaneous fat (r=0.18, P<0.01) in all subjects and correlated with TNF α(r=-0.16, P<0.05) and hsCRP (r=0.15, P<0.05) in subjects without MetS but not with MetS. With multiple linear regression analysis, these linear associations remained to be significant. The results of this study show that plasma resistin levels in humans were not associated with markers of insulin resistance, obesity or other determinants of the MetS

Simple sequence repeat variation in the Daphnia pulex genome

Background: Simple sequence repeats (SSRs) are highly variable features of all genomes. Their rapid evolution makes them useful for tracing the evolutionary history of populations and investigating patterns of selection and mutation across gnomes. The recently sequenced Daphnia pulex genome provides us with a valuable data set to study the mode and tempo of SSR evolution, without the inherent biases that accompany marker selection. Results: Here we catalogue SSR loci in the Daphnia pulex genome with repeated motif sizes of 1-100 nucleotides with a minimum of 3 perfect repeats. We then used whole genome shotgun reads to determine the average heterozygosity of each SSR type and the relationship that it has to repeat number, motif size, motif sequence, and distribution of SSR loci. We find that SSR heterozygosity is motif specific, and positively correlated with repeat number as well as motif size. For non-repeat unit polymorphisms, we identify a motif-dependent end-nucleotide polymorphism bias that may contribute to the patterns of abundance for specific homopolymers, dimers, and trimers. Our observations confirm the high frequency of multiple unit variation (multistep) at large microsatellite loci, and further show that the occurrence of multiple unit variation is dependent on both repeat number and motif size. Using the Daphnia pulex genetic map, we show a positive correlation between dimer and trimer frequency and recombination. Conclusions: This genome-wide analysis of SSR variation in Daphnia pulex indicates that several aspects of SSR variation are motif dependent and suggests that a combination of unit length variation and end repeat biased base substitution contribute to the unique spectrum of SSR repeat loci

Shifting patterns of natural variation in the nuclear genome of caenorhabditis elegans

<p>Abstract</p> <p>Background</p> <p>Genome wide analysis of variation within a species can reveal the evolution of fundamental biological processes such as mutation, recombination, and natural selection. We compare genome wide sequence differences between two independent isolates of the nematode <it>Caenorhabditis elegans </it>(CB4856 and CB4858) and the reference genome (N2).</p> <p>Results</p> <p>The base substitution pattern when comparing N2 against CB4858 reveals a transition over transversion bias (1.32:1) that is not present in CB4856. In CB4856, there is a significant bias in the direction of base substitution. The frequency of A or T bases in N2 that are G or C bases in CB4856 outnumber the opposite frequencies for transitions as well as transversions. These differences were not observed in the N2/CB4858 comparison. Similarly, we observed a strong bias for deletions over insertions in CB4856 (1.44: 1) that is not present in CB4858. In both CB4856 and CB4858, there is a significant correlation between SNP rate and recombination rate on the autosomes but not on the X chromosome. Furthermore, we identified numerous significant hotspots of variation in the CB4856-N2 comparison.</p> <p>In both CB4856 and CB4858, based on a measure of the strength of selection (k_a/k_s), all the chromosomes are under negative selection and in CB4856, there is no difference in the strength of natural selection in either the autosomes versus X or between any of the chromosomes. By contrast, in CB4858, k_a/k_svalues are smaller in the autosomes than in the X chromosome. In addition, in CB4858, k_a/k_svalues differ between chromosomes.</p> <p>Conclusions</p> <p>The clear bias of deletions over insertions in CB4856 suggests that either the CB4856 genome is becoming smaller or the N2 genome is getting larger. We hypothesize the hotspots found represent alleles that are shared between CB4856 and CB4858 but not N2. Because the k_a/k_sratio in the X chromosome is higher than the autosomes on average in CB4858, purifying selection is reduced on the X chromosome.</p

Proteomic characterization of HIV-modulated membrane receptors, kinases and signaling proteins involved in novel angiogenic pathways

<p>Abstract</p> <p>Background</p> <p>Kaposi's sarcoma (KS), hemangioma, and other angioproliferative diseases are highly prevalent in HIV-infected individuals. While KS is etiologically linked to the human herpesvirus-8 (HHV8) infection, HIV-patients without HHV-8 and those infected with unrelated viruses also develop angiopathies. Further, HIV-Tat can activate protein-tyrosine-kinase (PTK-activity) of the vascular endothelial growth factor receptor involved in stimulating angiogenic processes. However, Tat by itself or HHV8-genes alone cannot induce angiogenesis <it>in vivo </it>unless specific proteins/enzymes are produced synchronously by different cell-types. We therefore tested a hypothesis that <it>chronic </it>HIV-<it>replication in non-endothelial cells </it>may produce novel factors that provoke angiogenic pathways.</p> <p>Methods</p> <p>Genome-wide proteins from HIV-infected and uninfected T-lymphocytes were tested by subtractive proteomics analyses at various stages of virus and cell growth <it>in vitro </it>over a period of two years. Several thousand differentially regulated proteins were identified by mass spectrometry (MS) and >200 proteins were confirmed in multiple gels. Each protein was scrutinized extensively by protein-interaction-pathways, bioinformatics, and statistical analyses.</p> <p>Results</p> <p>By functional categorization, 31 proteins were identified to be associated with various signaling events involved in angiogenesis. 88% proteins were located in the plasma membrane or extracellular matrix and >90% were found to be essential for regeneration, neovascularization and angiogenic processes during embryonic development.</p> <p>Conclusion</p> <p>Chronic HIV-infection of T-cells produces membrane receptor-PTKs, serine-threonine kinases, growth factors, adhesion molecules and many diffusible signaling proteins that have not been previously reported in HIV-infected cells. Each protein has been associated with endothelial cell-growth, morphogenesis, sprouting, microvessel-formation and other biological processes involved in angiogenesis (p = 10^-4to 10^-12). Bioinformatics analyses suggest that overproduction of PTKs and other kinases in HIV-infected cells has <it>suppressed </it>VEGF/VEGFR-PTK expression and promoted <it>VEGFR-independent </it>pathways. This unique mechanism is similar to that observed in neovascularization and angiogenesis during embryogenesis. Validation of clinically relevant proteins by gene-silencing and translational studies <it>in vivo </it>would identify specific targets that can be used for early diagnosis of angiogenic disorders and future development of inhibitors of angiopathies. This is the first comprehensive study to demonstrate that HIV-infection alone, without any co-infection or treatment, can induce numerous "embryonic" proteins and kinases capable of generating novel <it>VEGF-independent </it>angiogenic pathways.</p

Phospholipase D signaling: orchestration by PIP2 and small GTPases

Hydrolysis of phosphatidylcholine by phospholipase D (PLD) leads to the generation of the versatile lipid second messenger, phosphatidic acid (PA), which is involved in fundamental cellular processes, including membrane trafficking, actin cytoskeleton remodeling, cell proliferation and cell survival. PLD activity can be dramatically stimulated by a large number of cell surface receptors and is elaborately regulated by intracellular factors, including protein kinase C isoforms, small GTPases of the ARF, Rho and Ras families and, particularly, by the phosphoinositide, phosphatidylinositol 4,5-bisphosphate (PIP2). PIP2 is well known as substrate for the generation of second messengers by phospholipase C, but is now also understood to recruit and/or activate a variety of actin regulatory proteins, ion channels and other signaling proteins, including PLD, by direct interaction. The synthesis of PIP2 by phosphoinositide 5-kinase (PIP5K) isoforms is tightly regulated by small GTPases and, interestingly, by PA as well, and the concerted formation of PIP2 and PA has been shown to mediate receptor-regulated cellular events. This review highlights the regulation of PLD by membrane receptors, and describes how the close encounter of PLD and PIP5K isoforms with small GTPases permits the execution of specific cellular functions

Search for new physics with dijet angular distributions in proton-proton collisions at root S = 13 TeV

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